Human-caused disturbance increasingly degrade global biodiversity and one of the main drivers is urbanization. It is well-known urbanization influence abundance and distribution of species. However evolutionary effect of urbanization is only beginning to be understood and no study examined rapid morphological change caused by biotic change in urban environment. In general, urban environment has less predator than natural habitat hence anti-predator defence of prey in urban environment could get weak because of the release from strong predation pressure. Here, we compared morphological traits (parotoid gland size, mouth width, tibia length, weight) of invasive cane toad in urban (Townsville) and forest (20 km away from the city), which was introduced to Australia in 1935. We found parotoid gland, which work as strong anti-predator defence, of urban toad were significantly smaller than those of forest toad, while there were no significant differences in other traits. These results suggest anti-predator defence of urban toad is likely to get weak compared with forest toad within approximately 80 years. Moreover, urban area is not isolated from forest area hence there may be gene flow amongst the populations. Our result may suggest that selective effect caused by urbanization overwhelms the effect of gene flow facilitating homogenization.